Changes in free cytoplasmic calcium (Ca2+) levels as measured by the fluorescent Ca2+ indicator Quin-2, release of inositol phosphates from phosphatidyl inositol (PI response) and histamine release were examined in rat basophil (2H3) cells. Exposure to antigen, Concanavalin A and Ca2+ ionophores resulted in a rapid increase in cytosol Ca2+ levels and over the course of 30 min release of histamine and inositol phosphates. The cytosol Ca2+ levels reached a maximum within 2 to 3 min and then declined slowly over the course of 30 min as the cells became refractory. None of these responses were observed in the absence of calcium but all were evoked upon subsequent addition of Ca2+. The Ca2+ and PI responses required the continued presenceof external Ca2+ and stimulant. The addition, for example, of EGTA after antigen had been added led to rapid abrogation of the Ca2+ response which could be restored immediately by addition of excess Ca2+ ions. All 3 responses could be blocked by methoxyverapanil, lanthanum salts and metabolic inhibitors, whereas zinc salts selectively blocked histamine release presumably by acting at intracellular sites. The manner in which responses varied with concentrations of stimulant and inhibitor indicated that histamine release was dependent on Ca2+ flux and ligand-IgE receptor aggregation whereas PI response appeared to be an independent process triggered by receptor aggregation.